Alpha-1 antitrypsin deficiency is an inherited, genetic condition characterized by reduced serum levels of alpha-1 antitrypsin (AAT) and increased risk of developing emphysema and liver disease, which affects up to 100,000 individuals in the United States. The only currently approved therapy for this orphan disease is weekly intravenous infusions of AAT protein that has been purified from normal plasma donors. The current therapy is in short supply, resulting in low physician motivation to accurately diagnose the vast majority of patients, since not all of the minority of patients who are currently diagnosed are able to receive treatment. The long-term objective of the research proposed in this application is to develop and bring to market a novel, gene therapy product for the treatment of alpha-1 antitrypsin deficiency, using a recombinant adeno-associated virus (rAAV) serotype 1 vector expressing the human AAT gene (rAAV1-CB-hAAT). rAAV vectors are uniquely suitable for in vivo gene therapy because they are non-toxic, highly efficient at transducing a wide variety of non-dividing cell types, and persist for long periods, primarily in episomal form, resulting in long-term expression of the transgene. The specific aim of this research proposal is to perform a Phase 2 clinical trial of rAAV1-CB-hAAT. The study is a non-randomized, open-label, multi-center, sequential, three-arm, Phase 2 clinical trial evaluating the safety and efficacy of administration of a rAAV1-CB-hAAT vector administered by intramuscular (IM) injection. Each participant will receive rAAV1-CB-hAAT on a single occasion. Three groups of three subjects each will receive rAAV1-CB-hAAT at dosage levels of 6 x 1011 vector genome (vg)/kg, 1.9 x 1012 vg/kg or 6 x 1012 vg/kg by IM injection. Subjects in group 1 will receive a total of 10 IM injections distributed across a single muscle site, subjects in group 2 will receive a total of 32 IM injections distributed across three muscle sites, and subjects in group 3 will receive 100 IM injections distributed across10 muscle sites. Each injection will be given in a volume of 1.35 mL, at the appropriate vector concentration to achieve the desired total vector dose, and the injection density at each administration site (nine IM injections per 4 cm2 skin surface area) will be the same as the injection density that was well tolerated in a previous Phase 1 clinical trial with rAAV1-CB-hAAT. Safety will be monitored by evaluation of adverse events, hematology and clinical chemistry parameters, histological examination of muscle biopsies, and measurement of serum antibodies to AAT. Efficacy will be measured by evaluation of serum concentrations of M-specific AAT and total AAT and serum AAT phenotype determined on isoelectric focusing gels. Additional information to be collected will include presence of the vector in blood or semen, changes in serum anti-AAV antibody titers, and changes in T cell responses to AAV and AAT. Results from this Phase 2 trial will be used to develop a Phase 3 study designed to satisfy the requirements for licensure of a gene therapy treatment for this orphan disease.